ORIGINAL RESEARCH
Engineering a recombinant Herpesvirus saimiri strain by co-culturing transfected and permissive cells
1 Moscow Institute of Physics and Technology, Dolgoprudny, Russia
2 Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, Moscow, Russia
Correspondence should be addressed: Stepan P. Chumakov
Miklouho-Maclay, 16/10, Moscow, 117997; moc.liamg@lukhtah
Funding: this study was supported by the Ministry of Science and Higher Education of the Russian Federation (Project ID RFMEFI60418X0205).
Acknowledgement: we thank to the Center for Precision Genome Editing and Genetic Technologies for Biomedicine (Moscow) for the genetic research methods.
Author contribution: Hamad A — manipulations on cell cultures, molecular cloning; Chumakov SP — study plan; manipulations on viral stocks, transfection, titration, data analysis, manuscript preparation.
Recombinant herpesviruses can be used as oncolytic therapeutic agents and high packaging capacity vectors for delivering expression cassettes into the cell. Herpesvirus saimiri is a gamma-herpesvirus that normally infects squirrel monkeys but also has a unique ability to infect and immortalize human lymphocytes while allowing them to retain their mature phenotype and functional activity. Recombination of the Herpesvirus saimiri genome in permissive cells is impeded by its resistance to chemical transfection and electroporation. The aim of this study was to develop an effective method for incorporating expression cassettes into the genome of Herpesvirus saimiri without having to transfect a permissive cell culture. Transfected HEK-293T cells expressing glycoproteins of the measles virus vaccine strain were co-cultured with permissive OMK cells infected with Herpesvirus saimiri. Cell fusion and formation of syncytia stimulated recombination between the viral genome and the expression cassette; this allowed us to obtain a recombinant Herpesvirus saimiri variant without chemical transfection in permissive cells. The genetically modified virus expressed a selectable marker and retained its ability to persist in the cell in the latent state; it also caused immortalization of primary lymphoid cells. The proposed approach allows engineering recombinant Herpesvirus saimiri strains carrying a variety of expression cassettes in its genome.
Keywords: CAR, chimeric antigen receptor, herpesvirus, viral vector, recombinant strain